Clostridium tetani

Tetanus- C/F

1. Clostridium tetani Dr V S Vatkar Asso Prof Microbiology Department D Y Patil Medical College, Kolhapur

2. Causative organism of tetanus Known since early time, described by Hippocrates & Aretaeus . Kitasato (1889): isolated it in pure culture and reproduced the disease in animals Widely distributed in soil and in intestine of animals & humans Other sources: street & hospital dust, cotton wool, plaster of Paris, bandages, catgut, talc, wall plaster, clothing etc

3. morphology Gram + ve , slender bacilli 4-8 by 0.5 µm size Arranged singly or in chains Spore bearing, spherical, terminal & bulging ( drum stick appearance ) Non-capsulated, motile- peritrichate flagella

4. Cultural characteristics Obligate anaerobes Opt temp: 37o C, pH- 7.4 Grows on ordinary media Blood agar: swarming, α hemolysis Deep agar shake culture: fluffy balls Gelatine stab culture: fir tree type growth with liquefaction RCM: turbidity, gas formation, meat pieces turns black

5. Colonies on blood agar

6. Biochemical reactions Indol + ve MR – VP: negative H2S: not formed Feeble proteolytic Does not ferment sugars

7. resistance Spores : more resistant to heat Killed by boiling 10-15 min, but some may resist boiling up to 3 hrs Autoclaving 121 0 C for 20 min : recommended Survive in soil: years & resistant to most antiseptics Not destroyed by 5% phenol, 0.1% mercuric chloride Iodine 1%, H2O2 kill spores within few hours

8. classification 10 serological types: based on agglutination (type I – X) Type IV : non flagellated strain All types produce same type of toxin & is neutralized by anti toxin produced against any one type TOXINS : Hemolysin ( Tetanolysin ) & a powerful neurotoxin: Tetanospasmin

9. Tetanolysin : heat labile, oxygen labile, no relevant role in pathogenesis of tetanus TETANOSPASMIN : toxin responsible for tetanus, oxygen stable, plasmid coded, Inactivated by heat at 65 0 C for 5 min Toxoided spontaneously or in presence of low conc formaldehyde

10. Tetanospasmin Good antigenic, neutralizes with specific anti toxin Simple protein, made up of single polypeptide chain Heterodimer consist of : heavy chain, light chain joined by disulphide bond MLD: for human, 130 nanograms Horse : more susceptible, guinea pigs, mice, goats & rabbits : in descending order, birds & reptiles: more resistant.

11. pathogenesis Little invasive power Washed spores : injected in experimental animals do not germinate & destroyed by phagocytosis Germination & toxin production occur only in favourable conditions like reduced OR potential, devitalized tissues, foreign bodies or concurrent infection Toxin: mainly absorbed by motor nerve endings & transported to CNS

12. Mechanism of action Resembles like strychnine Blocks synaptic inhibition of spinal cord (inhibitory terminals that uses glycine & GABA as neurotransmitters. Toxin acts pre synaptically (Strychnine acts post synaptically ) Abolition of spinal inhibition causes uncontrolled spread of impulse initiated anywhere in CNS Causes muscle rigidity & spasm (due to simultaneous contraction of agonist & antagonist muscles, in absence of inhibition)

13. Tetanus toxin blocks synaptic inhibition At inhibitory terminals that use GLYCINE & GABA neurotransmitters Toxin acts pre synaptically Abolition of spinal inhibition Uncontrolled spread of impulses Initiated anywhere in CNS Muscle rigidity & spasms due to Simultaneous contraction of agonist & antagonist, absence of inhibitory responce

14. tetanus Tonic muscular spasm Followed by injury, especially puncture wounds Rarely followed by surgery Sometimes may be due to suppurative foci like otitis media Septic abortion Unhygienic practices; application of cow dung on umbilical cord Unsterile injections

15. Incubation period: variable, 2 days – several weeks (6-12 days) Site & nature of wound Dose & toxigenesity of organism Immune status of the patient I P: short : prognosis: grave High mortality rate, 80-90% Tetanus neonatorum & uterine tetanus : high fatality rate

16. Common in developing countries: warm climate, rural areas: soil ,fertile & highly cultivated Unhygienic practices Universal immunization of infants & expected mothers: reduced incidence of tetanus, especially in rural areas

17. Laboratory diagnosis Always on clinical ground Lab tests: help for confirmation MICROSCOPY : unreliable, demonstration of drumstick bacilli in the wound itself is not diagnostic, Cl tetani, Cl tetanomorphum , Cl sphenoides : morphologically similar CULTURE : diagnostic, specimen should be from the deeper part of the wound. Plated on blood agar, incubated anaerobically, swarming seen

18. culture Material : also inoculated into 3 bottles of RCM 1 st bottle: heated to 80 0 C for 15min 2 nd bottle: heated to 80 0 C for 5 min 3 rd bottle: kept unheated Purpose of heating for different period : to kill vegetative bacteria, undamaged tetanus spores which may be heat resistant Bottles : incubated at 37 0 C & subcultured on blood agar plate daily for next 4 days Polymyxin B blood agar : selective medium

19. In vitro toxigenecity testing Blood agar containing 4% agar: tetanus antitoxin (1500 Units/ml): spared on 1/2 plate & test strain is inoculated on both halves, incubated anaerobically for 2 days. Toxigenic Cl.tetani : hemolysis around colonies on half plate without antitoxin & no hemolysis on half : plated with antitoxin

20. In vivo toxigenecity test Tested in animals: 2 mice 2-4 days old cooked meat culture (0.2 ml) is injected in root of tail of both mice One mouse is protected with tetanus antitoxin (1000 units) , other is unprotected Symptoms develop within 12-24 hrs , begins with stiffness of tail, rigidity to leg on inoculated side then opposite leg, trunk & forelimb (ascending tetanus) Animal dies within 2 days

21. prophylaxis Surgical prophylaxis : remove the foreign body, necrotic tissue, blood clot to prevent anaerobic environment Radical excision: depending on the type of wound Antibiotics : inhibit pyogenic bacteria & tetanus bacilli Long lasting Penicillin : drug of choice Erythromycin : 500 mg b d for 5 days Bacitracin / neomycin : applied locally

22. immunization Passive immunization : Tetanus antitoxin: injections Anti tetanus serum (ATS): prepared from hyperimmune horse, 1500 units s/c or i/m , given to non immune person after injury Reduces incidence & prolongs incubation period of tetanus Two disadvantages: heterologus serum: immune elimination & hypersensitivity

23. Half life of ATS: 7 days but in previously received person: eliminates quickly (combines with previously existing antibodies) Hypersensitivity: mild local reaction to serum sickness sometimes fatal anaphylaxis Human antitetanus immunoglobulins (TIG): small doses,250 units, longer half life(3-5 weeks), prepared from immunization human volunteers, availability limited

24. Active immunization Toxoided form : plain toxoid or adsorbed on aluminium hydroxide or phosphate Adsorbed toxoid : more antigenic Given along with diphtheria & pertussis (DPT vaccine) as triple vaccine Deep i/m at an interval of 4-6 weeks, first 2 doses, starting at the age of 1 & ½ month, 3 rd dose after 6 month or recommended in immunization schedule

25. Booster dose: 10 yrs Too frequent inj of T T should be avoided: hypersensitivity reactions may occur occasionally COMBINED IMMUNIZATION : given to non immunized person TIG inj at one site & first dose of TT 2 nd & 3 rd dose of TT at interval of 1 month

26. treatment Quite environment Controlling spasm: sedatives, muscle relaxtants Maintain airway: tracheostomy, + ve pressure ventilation Human TIG 10,000 IU, slow IV infusion, if needed repeat 5000 IU later Antibiotics: penicillin, metronidazole for wk Patient recover from tetanus : give full course of TT(active immunization)